Anti col1a1 antibody

We applied the Total Protein Extraction [Cat. no. C006225-0050, Sangon Biotech (Shanghai) Co., Ltd.] to extract the total proteins of different cell groups. BCA method (Beyotime, Shanghai, China) was applied for protein concentration SDS-PAGE electrophoresis was taken to isolate the total protein with 50 µg total proteins per pore. After 2-h electrophoresis, the proteins were wet transferred to PVDF membranes, sealed with 5% skim milk powder (1 h) and incubated with primary antibodies of Anti-Bax antibody (ab32503, 1:1000, Abcam, USA), Anti-bcl2 antibody (ab59348, 1:1000, Abcam, USA), Anti-Cleaved Caspase3 antibody (ab2302, 1:1000, Abcam, USA), Anti-E-Cadherin antibody (ab40772, 1:1000, Abcam, USA), Anti-Vimentin antibody (ab92547, 1:1000, Abcam, USA), Anti-SNAIL antibody (ab53519, 1:1000, Abcam, USA), Anti-COL1A1 antibody (39,952, Cell Signaling Technology, USA), Anti-beta Actin antibody (ab8227, 1:1000, Abcam, USA) overnight at 4 ℃. The next morning, the membranes were rinsed with TBST and incubated at 37 ℃ with the addition of horseradish peroxidase-labeled Goat Anti-rabbit (ab6721, 1:300, Abcam, USA) (1 h). The protein bands were exposed by Chemistar™ High-sig ECL Western Blotting Substrate (TANON SCIENCE & TECHONLOGY CO., Shanghai, China). The experiment was repeated three times.

Histological and immunohistochemical sections were prepared [27 (link)] and stained with hematoxylin and eosin (HE) and Masson’s trichrome (MT) according to standard protocols. For IHC, antigens were activated using a Histofine simple stain kit (Nichirei Biosciences, Tokyo, Japan), and the sections were stained with anti-COL1A1 antibody (Cell Signaling Technology, 72026), anti-α-SMA antibody (Cell Signaling Technology, 19245), anti-ZEB1 antibody (Cell Signaling Technology, 70512), anti-SNAIL + SLUG antibody (Abcam, ab180714), and anti-Lamin B1 antibody (Synaptic Systems GmbH, Göttingen, Germany, HS-404013). TUNEL staining was performed using the In Situ Cell Death Detection Kit (Sigma Aldrich, 1684795).